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Standard Operating Procedures

Silver Salt Analysis

Introduction

This document describes how to set up and run nitrate and sulfate for Δ17O on the Lorax, ThermoFinnigan MAT 253. Nitrate and sulfate samples (in the form of silver precipitate in silver or gold capsules, respectively) are dropped by an autosampler into a hot quartz pyrolysis reactor using a Thermal Conversion Elemental Analyzer (TCEA).  A helium carrier stream moves pyrolysis products through a liquid nitrogen trap where byproducts other than O2 are frozen.  The O2 product then continues through a gas chromatograph and either a Conflo or Gas Bench interface.  The Conflo is used for samples of O2 > 2 μmol; the Gas Bench cryofocuses O2, enabling analysis of smaller samples (50-1000 nmol).  Make sure the sample is <1000nmol when using Gas Bench mode. Helium continues to move O2 into the mass spectrometer for m/z 32, 33, and 34 measurements.

This general plumbing diagram and this gasbench valco valve diagram may be helpful.

Nomenclature

Liquid Nitrogen Traps:

  1. TCEA pre-cleanup trap = ¼” tubing filled with 5A molecular sieve submerged in liquid nitrogen to scrub O2 from helium stream before entering the reactor.
  2. TCEA post-cleanup trap = 1/16” tubing submerged in liquid nitrogen to remove post-pyrolysis byproducts (NOx and SO2, mostly)
  3. Big trap or Concentrating trap = 1/16” tubing molecular sieve trap on Gas Bench and referred to as Trap 2 in software
  4. Little trap or Cryofocusing trap = capillary molecular sieve trap on Gas Bench and referred to as Trap 1 in software

Valves:

  1. Autosampler Purge Valve = three-way valve, top of autosampler carousel to either evacuate the autosampler or allow venting of helium
  2. He Purge Valve = gold cylinder, side of autosampler carousel to allow helium into the carousel / autosampler for purging
  3. Isolation Valve = knob, between autosampler carousel and reactor column to isolate the autosampler from the reactor
  4. Vent Valve  = 3-way valve on rear top of TCEA to vent after installing a fresh column (vent) or to allow helium stream to pass on past the TCEA (run)
  5. Method Valve = 4-way valve on rear top of TCEA to choose Gas Bench or Conflo
  6. Auxillary He Valve = 2-way valve next to Method Valve to allow helium flow to both methods (Gas Bench and Conflo)
  7. Valco Valve = 8-way valve, toggle in Gas Bench software; not used in Conflo method
  8. Needle Valves = IRMS inlet; top valve is from Gas Bench, side valve is from ConFlo

Instrument Preparation

  1. Make sure the pyrolysis column is clean and ready for another set of samples (see Pyrolysis Column below).
  2. Bake what needs to be baked:
    1. To bake the GC contained inside the TCEA (for Conflo mode), use the JUMO iTRON 16 labeled GC on the front of the TCEA, press ‘P’ until you see ‘t 0’, then press the ‘up arrow’. This will ramp the GC within the TCEA to 300 *C for 10 hours. Or if you don’t see ‘t 0’, press ‘P’ until you see ‘SP’ (not ‘SPΓ’), then press the ‘up arrow’ to set the temperature to 300°C. But this method will not set the backing time for 10 hours, and you will need to set the temperature to 35°C manually in the same way after 10 hours. 
    2. Use the kitchen timer located above the TCEA (look at the ceiling) to bake the TCEA clean-up traps for 10 hours. Make sure the controller located to the left of the TCEA and labeled ‘TCEA He cleanup trap’ is set to ‘HI’.
    3. To bake the GC inside the Gas Bench, use the JUMO iTRON 16 on the side of the Gas Bench, press ‘P’ until you see ‘t 0’, then press the ‘up arrow’. This will ramp the GC within the TCEA to 200 *C for 10 hours. Or use the ‘SP’ method described above in 2a.
    4. To bake the Gas Bench Traps (Big Trap and Little Trap), install the heater box and a lab jack to enclose the traps. Use the same kitchen timer as above set for 10 hours and make sure the controller to the left of the TCEA labeled ‘Gas Bench Traps Box’ is set to 2.5.
  3. If you are switching between Conflo and Gas Bench, close the Needle Valve that is open BEFORE opening the other one.
    1. To close a Needle Valve, make sure the vacuum is visible to you while you stand at the valves. Close the valve ONLY as much as needed. YOU CAN CLOSE IT TOO MUCH AND RUIN THE VALVE. Watch the vacuum. When it reaches <3e-8 mbar, stop. WAIT. If it doesn’t appear to be dropping, GENTLY close it some more. If the vacuum responds, WAIT and repeat.
  4. The appropriate Needle Valve should be open the day before you intend to run.
    1. Make certain the dilution is ON and make the vacuum visible while you are standing at the Needle Valve. 
    2. VERY VERY VERY SLOOOOOWWWWLY open the appropriate Needle Valve watching the vacuum. You will see the vacuum rise from <2e-8 mbar to ~9.7e-7 mbar. 
    3. If you have just opened a Needle Valve that was previously closed, run an 8 hour bake-out sequence while in IsoDat ‘Conflo’ mode.

Pyrolysis Column

  1. You will either need to replace the pyrolysis tube or clean out the previous runs samples before starting a new run. 
  2. Turn the He dilution ON if running Conflo mode. Change the Valco Valve to Inject if running in Gas Bench mode.
  3. Turn Vent Valve to vent. 
  4. Use gloves and protective eye-wear. Unscrew the pyrolysis column starting at the top by loosening the nut and setting the autosampler towards the back of the TCEA.
  5. Unscrew the bottom fitting. 
  6. Remove pyrolysis tube by pushing the bottom up and grabbing the o-ring and place in the pyrex baking dish.
  7. Select a new pyrolysis tube or clean out the existing one with the wire-tipped wand. If the pyrolysis tube has never been used, you can test it for leaks using the fittings in the bag labeled “Reactor Vacuum Test” in the cabinet next to the Lorax computer. Put the rubber stopper in one end, attach the other and connect the glass tube to a vacuum line (a couple are next door). If the pyrolysis tube pumps down, it is ready to go.
  8. Place rubber o-ring on top of the new pyrolysis column about 5 mm from the top.  Use compressed air (red tube above TCEA) to clean any dust off ferrule. Slide pyrolysis tube into TCEA. Start at the top of the TCEA. While bearing the weight of the autosampler, screw on the top column nut and use a spanner to tighten an additional ¼ turn. Now move to the bottom of the column. Use compressed air to cool the bottom of the tube that just went through the furnace. Slide fitting onto the pyrolysis tube as high as it can go and ensure you have the proper order of the nut, back of ferrule, and front of ferrule and then finger tighten. Use wrenches to tighten an additional ¼-1/2 turn. DO NOT OVERTIGHTEN.
  9. The top rubber ferrule will compress over time, but will recover if allowed to rest for a few days – swap it with another rubber ferrule when it looks compressed or is leaking.  The bottom Teflon ferrules compress over multiple uses.  Once they start leaking (or get dirty), they must be replaced with new ferrules.  Teflon ferrules are either 0.9 inches or 23 mm in diameter.
  10. Use Leak detector set in “High” mode to check for leaks. Check around both top and bottom fittings as well as in grate in front of the TCEA and inside the heating block (accessed from the bottom). If leaks are present, repeat process of loading the pyrolysis tube without actually removing the column.
  11. Once leaks are eliminated, leave Vent Valve on vent for ~10 minutes before switching to run. 

Sample Loading

  1. Each sulfate run can contain a maximum of 10 samples in gold capsules or 8 samples in quartz capsules. Nitrate runs can have up to a full carousel (49 samples in folded/balled silver capsules).
  2. Isolate Autosampler by closing Isolation Valve. Arrow on side of valve should point towards you and you won’t be able to see down inside the reactor.
  3. Turn the Autosampler Purge Valve clockwise to vent pressure in the Autosampler. The valve handle will point towards computer.
  4. Loosen the three clamps on the autosampler (with spanner if needed). Open the top, and use compressed air line to clean out any bits of dust.
  5. Load samples into slots. Close the lid, and hand tighten each clamp, alternating between knobs to ensure even pressure.
  6. Turn Autosampler Purge Valve 180° counter-clockwise to evacuate the carousel. While it’s evacuating, tighten the lid with your fingers again.
  7. After 3-5 minutes, turn Autosampler Purge Valve to point straight back (off position).
  8. Slowly open He Purge Valve. Monitor pressure on the front of TCEA until it levels off (Purge=1.5 bars and Carrier=1.5 bars).
  9. Use leak detector to check for autosampler leaks as done for the reactor. 
  10. If no leaks are found, slowly turn the Autosampler Purge Valve clockwise until the helium vents and the pressure gauges drop. Turn the Autosampler Purge Valve countclockwise to the straight back position. Once the pressure gauges read 1.5 bars, vent the helium again and repeat 10-12 times. Monitor the O2 background. Then slowly close He Purge Valve. If in Conflo mode, and if there is a leak on the pyrolysis column (even very small leak), then the O2 background will increase dramatically (>100 mV or even 2000 mV), and you should use a new pyrolysis column.
  11. Make sure the dilution is ON if in Conflo mode or Valco Valve is in inject if in Gas Bench mode and then open the Isolation Valve. 
  12. If in Conflo mode, monitor the O2 background signal for ~5 minutes. If the background has not increased dramatically since opening the Isolation Valve, turn the dilution off.
  13. Freeze the liquid nitrogen trap.
  14. Vent the helium using the Autosampler Purge Valve as above but this time the pressure gauges will NOT drop. Make certain to return the Autosampler Purge Valve to the straight back position BEFORE the hissing of helium stops. Repeat this venting 3-5 times.
  15. If in Conflo mode, monitor the O2 background signal for ~ 5 minutes. If it is not rising proceed to next section, otherwise repeat the above venting procedure 3-5 more times.
  16. Create a sequence using the IsoDat software (Acquisition or Workspace if something is running). For ConFlo mode, use the sequence template labeled Conflo_O2_Analysis_Template.seq. For Gas Bench, use GasBench_O2_Analysis_Template.seq. Record the amount of expected O2 for each sample (μmol or nmol), sample name, any other notes, and autosampler carousel slot. Runs in ConFlo have a “Zero” at the beginning and end of the run, as well as in between each sample. Make sure that Zero runs use method Conflo_O2_Zero.met and sample runs use method Conflo_O2_Sample.met. Runs in Gas Bench mode have a “Blank” at the beginning and end of the run, as well as in between each sample. Make sure the blanks use method GasBench_O2_Blk.met and samples use GasBench_O2_Sample.met (‘split-in’ or ‘split-out’, suggest using ‘split-out’ for samples amount between 200 and 1000 nmol).
  17. Save your sequence using the naming scheme YYMMDD_Conflo_RunName.seq for a conflo run or YYMMDD_GasBench_RunName.seq.

Daily Log

The lab uses a daily log for each instrument or preparation line to allow users a first glance at the readiness of the instrument. By comparing the current state of the instrument to historical states, you are more informed about the instrument and whether or not it is functioning properly and ready to run your samples.

Each daily log is web based and browser accessible. No link is provided here by design. Open the browser on the controlling computer and you should see at least two tabs already open. One tab is this SOP and the other is the daily log. If the browser has more than two tabs open, it may have additional SOPs. Use the bookmark toolbar as needed if tabs have been closed.

Work through each cell of the daily log. If you are uncertain where to find certain information, hover over the column header tip, denoted by a question mark (?).

Make certain to press the 'save to log' button when you are finished entering data.

You are welcome to make notes if you have observed something with or done something to the instrument and would like to document that information. Use the "insert note" link at the top of the daily log to make a note. You may enter notes at any time.

Running in conflo mode

  1. Ensure that all valves are in the proper orientation: 
    1. Autosampler Purge Valve closed
    2. Isolation Valve open
    3. He Purge Valve closed
    4. Mode Valve in Conflo mode
    5. Needle Valve open
    6. Top off TCEA cleanup trap with LN2. The LN2 in the trap will need to be topped off several times during the course of a day, so check periodically to ensure appropriate levels.
  2. Open your sequence in IsoDat Acquisition. If you are not running every row in the sequence, highlight the appropriate rows, and click the green “Start” button.
  3. Fill in the appropriate fields with folder/file names. Make sure that the export template selected is “Conflo.wk”, and that the export file type is comma separated values (.csv). Click ‘OK’ and your run will start.

Running in gas bench mode

  1. Ensure that all valves are in the proper orientation: 
    1. Autosampler purge valve closed
    2. Isolation valve open
    3. He purge valve closed
    4. Mode valve switched to Gas Bench mode
    5. needle valve open
    6. Top off TCEA cleanup trap with LN2 and fill a large blue dewar with LN2 and place below the cryofocusing traps. The LN2  level should be within ~2cm of the top of the dewar.  Again, check the LN2 periodically and top off as needed. 
  2. Run several blanks with the system setup as if you were running. Once there are three blanks in a row that are sufficiently small (Area all < 30 Vs for the split-in method and <4 Vs for the split-out method) and the same size, you may proceed with the run. If blank sizes are large, try re-purging autosampler, as this is the most likely source of the blank. 
  3. Open your sequence in IsoDat Aquistion. If you are not running every row in the sequence, highlight the appropriate rows, and click the green “Start” button. Fill in the appropriate fields with folder/file names. Make sure that the export template selected is “Conflo.wk”, and that the export file type is comma separated values (.csv). Click begin 

Post-run cleanup

  1. When a run is done (in either mode), turn the vent valve to “Vent” mode and thaw the TCEA cleanup trap by removing it from the LN2 trap and baking it on ‘HI’ for 5 minutes.
  2. Pour any leftover LN2 back into a storage dewar.
  3. Clean out the pyrolysis column as described above under Pyrolysis Column.
  4. Change the Vent Valve back to run mode.
  5. When running is Gas Bench mode, make sure the Valco valve is in “Inject” mode (toggle on the IsoDat control screen). Empty large dewar back into LN2 storage dewar. 

Data processing

Will need to refine this once we settle on a consistent way to handle data (i.e. what graphs we want, whether we want to do a script for conflo and gas bench or if one is sufficient, what corrections we want, etc.) 

There are Matlab scripts set up in order to correct samples and reduce the raw data file into a condensed form that is easier to use. After verifying that sample peaks look as expected (i.e. no secondary peaks/dips in background), copy the raw data file onto a jump drive. For both Conflo and Gas Bench, the file is found in the “ConFlo Results” folder, which can be found on the desktop of the Lorax computer. Your chosen folder name will be present here, and within it a folder labeled Excel. The file is inside this folder. 

Log in to one of the computers in the northwest corner of the IsoLab (labeled Southgoing Zax and Northgoing Zax). Both computers are linked to an external hard drive, so you can use either one. Copy your file into the appropriate subfolder within the Data folder, a shortcut to which is on the desktop. In the Data folder, double click on the Projects folder.  

If you only ran standards, click on the “Standards” folder, then on either “Sulfate” or “Nitrate” and finally “Raw”. Paste your file here. 

If you do not have a subfolder in “Projects” already, create one now. Create subfolders called “Raw” and “Reduced”. Paste file into the “Raw” folder.  

Open Matlab. Type “lorax” into the command line. A window will automatically appear. Locate and double click on your file. Matlab will run the raw file and select all sample and standard lines and export a reduced file that does not have any “zero” or extraneous data. Matlab will also automatically create a series of graphs plotting data against analysis number: Δ17O, yield, peak start time, and two proxies for peak shape (amplitude32/width32; amplitude32/area32). The script will also log all of the standards from a run into a separate archive file. The reduced file will appear in the “Reduced” folder and can be opened in either Matlab or Excel.  

Troubleshooting

ProblemPossible CauseSolution
Background levels are highAutosampler not purged fullyRe-purge autosampler
Background levels are still highGC/Cryo Traps retaining remnants of atmospheric airBake out GC Cryo Traps IRMS
Conflo - Sample peak timing delayedLeak somewhere (likely reactor)Fix Leak
Gas Bench - Sample peak shape off or Sample peak timing earlyCryo Traps saturated with atmospheric airBake out GC-Cryo Traps-IRMS
Gas Bench - Sample peak shape off-Tiny sample peakAnti-blank (production of Ag2O instead of O2 released?)Use Gold capsules or Try different temperatures
Sample peak too smallLeak on pyrolysis columnUse a new pyrolysis column

Reference materials

Nitrate:

Sulfate:

Last updated: 150206 by Qianjie Chen